SCF complexes are ubiquitin ligases that degrade a diverse group of substrates, and the F-box proteins contained in these complexes mediate substrate recognition. Fbw7 is an F-box protein that targets cyclin E, Myc, Notch and Jun for degradation. Fbw7 is frequently mutated in human cancers and functions as a tumor suppressor in mice. The mechanisms of tumor suppression by Fbw7 are largely unknown, but likely involve the oncogenic deregulation of its downstream targets. The overall goal of this application is to understand how Fbw7-loss contributes to the development of epithelial cancers, and it focuses on the role of cyclin E in Fbw7-associated cancer. The goal of the first Aim is to utilize adeno-associated virus gene targeting techniques to create Fbw7-null mutations in cultured epithelial cells. These cell models will be used to study the regulation of specific substrates and their roles in genetic instability. Because Fbw7-loss occurs frequently in human colon cancer, the second two Aims will utilize two complementary mouse models to study Fbw7 and cyclin E function in the colonic epithelium and their role in the development of colon cancer. In the second Aim, I will use a conditional-null Fbw7 strain to specifically inactivate Fbw7 function in the colonic epithelium. In the third Aim, I will utilize a knockin strain developed in our lab, in which cyclin E degradation by Fbw7 has been impaired by the mutation of a regulatory cyclin E phosphorylation site, to study deregulated cyclin E activity to the colonic epithelium. By comparing these two models, I will determine the role of Fbw7-loss and the importance of cyclin E deregulation in the development of colon cancer. Because both Fbw7 and cyclin E are intimately linked to the p53 pathway, these Aims will also study the interactions of the p53 and Fbw7 pathways in cell proliferation and transformation. The applicant's career goals are to establish an independent research program that integrates basic studies of cancer biology with his training in medical oncology. The proposed research will provide broad-based training that will provide the applicant with the skills needed to achieve these goals. Due to his expertise in the fields of cell cycle control and cancer biology, Bruce Clurman is ideally suited to supervise these efforts. Furthermore, the Fred Hutchinson Cancer Research Center is a world-renowned research environment that is dedicated to the training of career scientists. Relevance: The Fbw7 pathway is implicated in the development of many human cancers. Understanding the mechanisms that link the Fbw7 pathway to cancer will provide fundamental knowledge about the development of cancer. As such, it is a crucial step towards developing treatment strategies for tumors associated with this pathway.